Amino acid analysis by gas—liquid chromatography of N-heptafluorobutyryl isobutyl esters : Complete resolution using a support-coated open-tubular capillary column

Abstract

Amino acids have been separated by gas—liquid chromatography as their N-heptafluorobutyryl isobutyl esters. Complete resolution of derivatives of all the common amino acids has been achieved using a high-performance support-coated open-tubular capillary column. The analysis time was 30 min. Modifications to the derivatization procedure of MacKenzie and Tenashuk have been introduced. Acylation by heating at 150° was shown to be destructive; 110° has been selected for routine preparation. To obtain a volatile histidine derivative it has been found necessary to add an antioxidant and to heat samples with ethoxyformic anhydride prior to injection. Amino acid analysis of β-lactoglobulin after 6 N HCl digestion yielded results in good agreement with those obtained by the conventional ion-exchange method. The method has also been successfully applied to estimation of the different caseins in whole casein and in purified fractions by amino acid analysis of residues liberated by carboxypeptidase digestion.